Project Summary Precisely balanced proliferation and differentiation is required for development, growth, and repair of the eye. Imbalanced proliferation and differentiation is linked to neurodegenerative disorders as well as retinal cancers. This project is designed to uncover how cells in the growing zebrafish eye transition from proliferation to differentiation. Zebrafish develop rapidly and are particularly amenable to high-resolution imaging and genetic manipulations, allowing developmental events to be studied in real time and correlated with genetic, molecular and cellular changes. Three specific aims investigate how extrinsic input from growth factors and the underlying extracellular matrix influence cell proliferation and differentiation in the zebrafish retina. Aim 1: To uncover the cellular mechanisms downstream of Gdf6a-retinoic acid antagonism that maintain stem and progenitor cells in the growing zebrafish retina This aim tests the hypothesis that retinoic acid (RA) pathway activity favors symmetric, differentiating divisions in the growing retina. It employs genetic and pharmacological manipulations of RA signaling and time-resolved confocal imaging to analyze cell division plane orientation and map the composition of clones arising from single retinal progenitor cells. Aim 2: To explore how dorsal-ventral retinal patterning impacts eye growth and development This aim asks how dorsal signaling pathways regulated by Gdf6 guide changes in cell shape, size, orientation, and proliferative behavior within a stem cell niche in the zebrafish retina by employing genetic manipulations coupled with quantitative image analyses. Aim 3: To probe the relationship between a specialized basement membrane and stem-cell mediated growth in the zebrafish retina This aim builds on preliminary data linking changes in basement membrane organization with maintenance of retinal stem cells. It employs established methods and novel transgenic lines to test the hypothesis that deposition of Collagen 15a1b, establishes a specialized extracellular matrix that (1) anchors stem cells at the retinal periphery and (2) regulates their proliferation. Significance: The results from this project will impact our understanding of growth, repair, and regeneration in the retina and entire central nervous system. Specifically, the data will deepen our understanding of how extrinsic factors coordinate proliferation, differentiation, and cell survival. These data will also shed new light on retinal stem and progenitor cell behaviors that may be leveraged for treating neurodegenerative diseases and neuronal cancers.